The present invention relates to a polyene-specific glycosyltransferase derived from Pseudonocardia autotrophica. 
Polyene macrolides are a large family of natural products typically produced by soil actinomycetes. Polyene macrolides are usually biosynthesized by modular and large type I polyketide synthases (PKSs), followed by several steps of sequential post-PKS modifications. In the late stages, a cytochrome P450 oxidizes a methyl side chain to form an exocyclic carboxyl group, and a glycosyltransferase (GT) catalyzes addition of mycosamine, a deoxyaminosugar derived from GDP-d-mannose. Especially, glycosyltransferases are an important class of enzyme and are essential for the biosynthesis of glycosylated natural products because they catalyze the attachment of a sugar to an aglycone. These sugars are often essential for the pharmacological properties, including water solubility and/or the biological activity, of the compounds. Several complementary strategies, including semisynthesis, pathway engineering, and in vitro enzymatic glycosylation techniques, have emerged from recent studies as effective means of altering the natural product sugar structures.
The biosynthetic gene clusters for several polyene macrolides have been characterized. Most polyene antibiotics contain a single deoxyaminosugar attached to the macrolactones. Chemical modification studies have shown that adding sugar residues to polyenes can improve their pharmacological properties. Some polyenes naturally contain additional sugar residues attached to mycosamine. 67-121C, a dissacharide-modified aromatic heptaene, has been isolated from Actinoplanes caeruleus. The second sugar residue has been identified as GDP-d-mannose. It was dentified that the extending glycosyltransferase gene, pegA, catalyzed addition of a mannosyl residue to the mycosaminyl sugar during 67-121C biosynthesis. Another example, nystatin P1 with a disaccharide mycosamine-glucose was proposed by MS-MS analysis and a biosynthetic gene cluster identified in the Pseudonocardia P1 strain collected from Apterostigmaden tigerum garden worker ants. The second sugar residue has not been fully identified but is thought to be a hexose. The gene for the extending GT has been identified and was named nypY.
Most recently, the present inventors identified nystatin-like polyene (NPP) containing a disaccharide, mycosamine (α1-4)-N-acetyl-2-aminoglucose. Interestingly, NPP harboring a disaccharide moiety had more than 300-fold higher solubility and 10-fold lower hemolytic activity than nystatin, which contains only mycosamine. The additional N-acetyl-glucosamine increased the solubility of the polyene compound. However, the gene for this extending GT was not found in the main biosynthetic gene cluster.
Meanwhile, Korean patent publication No. 10-2010-0089370 discloses biosynthetic gene clusters for polyene derived from Pseudonocardia autotrophica and base sequence thereof, however does not mention a novel polyene-specific glycosyltransferase according to the present invention.
Accordingly, the present inventors identified the extending GT in NPP biosynthesis by draft genome sequencing and PCR-targeted gene disruption-complementation system then assessed the function of the coding gene in different polyene-producing strains.